The pathologic proteolytic activation of digestive zymogens within pancreatic acinar cells is a key step in the initiation of pancreatitis. The extracellular and intracellular mechanisms responsible for this activation remain unclear. Supraphysiologic concentrations (>10 fold than required for maximal secretion) of the physiologic ligand, cholecystokinin (CCK), given in vivo cause zymogen activation and pancreatitis. Similarly treated isolated acinar cells respond with zymogen activation and are injured. Factors that can sensitize the acinar cell to the pathologic effects of CCK might be relevant to the pathogenesis of acute pancreatitis. Several experimental studies suggest that CCK-activated pathways contribute to disease. These include pancreatitis induced by ischemia, bile salts, the CDE diet, and ethanol. We hypothesize that: 1) clinically relevant agents or conditions can either act alone or sensitize the acinar cell to the effects of CCK on zymogen activation, and 2) such activation will depend on Ca2+-dependent and independent mechanisms. We find that the following relevant factors sensitize the acinar cell to zymogen activation caused by physiologic concentrations of CCK: secretin, VLDL, and short chain fatty acids. With respect to alcohols, sensitization increased with chain length and deceased with branching. At sensitizing concentrations, alcohols caused no detectable changes in either cytosolic Ca2+ signaling or secretion. However, the sensitizing effects of secretin corresponded to effects on CCK induced Ca2+ signaling. Previous studies suggested that a low pH compartment mediated zymogen activation. We find that inhibitors of vacuolar (v) ATPase (bafinomycin and concanomycin) block CCK-induced zymogen activation. Using antibodies to subunits of vATPase, we observed translocation during CCK treatments. Thus, vATPase appears to be activated and contribute to zymogen activation. We plan to examine the cellular signaling pathways stimulated by these sensitizing agents with particular emphasis on Ca2+, cAMP, PKC, and vATPase. Further, we will examine the effects of the sensitizing agents on cell injury and the trafficking of active enzymes.